G protein-coupled receptors (GPCRs) form one of the largest protein families found in nature. To understand how these receptors function at a molecular level is the major focus of this research program. By using a combined molecular genetic/pharmacologic approach, the molecular mechanisms involved in GPCR folding and assembly, GPCR activation, and ligand/receptor/G protein interactions were explored. For these studies, different muscarinic acetylcholine and vasopressin receptor subtypes served as model systems. Studies with "split" m3 muscarinic receptors showed that GPCRs can be assembled from multiple independently stable building blocks. These data prompted us to speculate that the reconstitution of functional receptor complexes in vivo by protein fragments may represent a novel strategy in the treatment of diseases caused by inactivating mutations in distinct GPCRs. Consistent with this notion, we could recently demonstrate that truncated V2 vasopressin receptors known to be responsible for X-linked nephrogenic diabetes insipidus can be functionally rescued (in cultured cells) by coexpression with a C- terminal V2 receptor fragment missing in the mutant receptors. To elucidate the molecular basis of receptor/G protein coupling selectivity, we employed a novel experimental approach involving the coexpression of mutant GPCRs with hybrid Galpha subunits. Using this strategy, we could identify, for the first time, a specific contact site between a short segment of a GPCR (m2 muscarinic receptor) and a short sequence on Galpha (C-terminus of Galphai/o). We could demonstrate that this interaction is required and sufficient for receptor-mediated activation of Gi/o. More recently, we started to explore the structural elements determining the coupling selectivity of GPCRs activated by peptide ligands. Functional studies with hybrid V1a/V2 vasopressin peptide receptors showed that the differential G protein coupling profiles of individual members of a structurally closely related peptide receptor family can be specified by different single intracellular receptor domains. The molecular nature of the agonist-induced structural changes in GPCRs (resulting in receptor activation) remains unknown at present. By employing a novel insertion mutagenesis strategy (model system: m2 muscarinic receptor), we could demonstrate that agonist-induced receptor activation involves a relative movement of the sixth transmembrane helix towards the cytoplasm.